Figure 3.

Msn2/4 positively regulate ATG39 promoter activity through the STRE elements. (A) Two putative STRE elements in ATG39 promoter region. (B) Effects of MSN4 overexpression on expression of the P_ATG39-GFP reporter mutated in putative STRE elements. Wild-type (WT) strains harboring the P_ATG39-GFP reporter mutated in putative STRE elements and the plasmids with or without the MSN4 gene were grown at 25 °C until exponential phase, and then harvested. The GFP mRNA levels were quantified by qRT-PCR analysis, and relative mRNA levels were calculated using ACT1 mRNA. The values are plotted as the fold change from cells harboring the plasmids without the MSN4 gene. The data show mean ± SEM (n = 4). NS, not statistically significant (P > 0.05), as determined by Student’s t-test. (C,D) Effects of mutations in putative STRE elements on ATG39 upregulation induced by ER stress and nitrogen starvation. Wild-type (WT) cells harboring wild-type or mutated P_ATG39-GFP reporters were grown at 25 °C until exponential phase and treated with 3 μg/ml tunicamycin (TM) (C) or incubated under nitrogen-starved conditions (D) for the indicated time. The GFP mRNA levels were quantified by qRT-PCR analysis, and relative mRNA levels were calculated using ACT1 mRNA. The values are plotted as the fold change from wild-type cells harboring the integration which expresses GFP under the control of wild-type ATG39 promoter at the time of TM addition or nitrogen removal. The data show mean ± SEM (n = 3). **P < 0.01 as determined by Student’s t-test.